Development of a rapid enzyme immunoassay for the detection of retinol-binding protein

Abstract

Retinol-binding protein (RBP) was chosen as a surrogate marker for retinol because of the close correspondence between retinol and RBP. To meet the need for rapid, cost-effective determination of vitamin A status in populations, a quantitative enzyme immunoassay (EIA) for detection of RBP was developed. The resulting RBP EIA, a competitive assay, uses RBP adsorbed to microtest strip wells to compete with RBP in serum. The assay takes approximately 40 min. With a reference panel of sera, test accuracy was found to be within 4% of expected values through the calibrated range of 0.48-1.92 micro mol RBP/L (10-40 micro g RBP/mL). Intraassay and interassay variability averaged 6.7% and 8.9%, respectively. Specificity testing showed no interference from other serum proteins, prealbumin, rheumatoid factor, bilirubin, estrogen, or C-reactive protein. The RBP EIA provided linear results between 0.43 and 1.80 micro mol RBP/L (9 and 38 micro g RBP/mL). Preliminary laboratory evaluations indicated that the RBP EIA correlates well with radial immunodiffusion for RBP and with HPLC for retinol, the current reference standard. A field evaluation in a population at risk for vitamin A deficiency (VAD) resulted in close correlation between RBP EIA measures and retinol measures by HPLC (R(2) = 0.82). The RBP EIA is as reliable in estimating VAD as is HPLC retinol. After successful validations, the test should enable public health authorities to rapidly monitor VAD and track vitamin A status in populations.